Genetic marker for establishing and identifying dairy cattle molecule system spectrum and application of genetic marker
A genetic marker and molecular pedigree technology, which is applied in the determination/examination of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc. , to achieve good application prospects and economic benefits, high detection efficiency, and optimized detection costs.
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Embodiment 1
[0028] Example 1. Selection and determination of microsatellite sites
[0029] 1. The microsatellite marker provided by the present invention consists of 8 sites, namely TGLA227, TGLA122, BMC1207, BM103, INRA037, INRA134, BP7 and MB026; the basic information of the 8 sites is shown in Table 4 below:
[0030] Table 4 Basic information of loci
[0031]
[0032]
[0033] The nucleotide sequence of the above primer sequence is shown in SEQ NO.1-16;
[0034] 2. Microsatellite locus genotype detection
[0035] Using single-plex fluorescent PCR combined with ABI3730 automatic sequencer fluorescence electrophoresis technology, the genotypes of PCR products at 8 sites in 300 dairy cows and 9 bulls were detected. The fluorescently labeled PCR amplification products were subjected to capillary electrophoresis, and the genotype peak was detected by ABI3730 sequencer, and the microsatellite genotype was determined by Gene Mapper V3.0 software combined with Gene Marker V1.75 softwar...
Embodiment 2
[0064] Example 2 Pedigree Accuracy Verification
[0065] In order to verify the accuracy and reliability of 8 microsatellite loci for paternity testing in dairy cow populations, this example carried out paternity testing on 8 pairs of father-son cows with paper pedigree records and father-son relationships. Genomic DNA of 8 parent-child pair cattle and 1 unrelated father cattle were extracted, and the genotypes and gene frequencies of 8 microsatellite loci were analyzed. The following results are shown in Table 10. Use the Parentage analyzes is module of the Analyze is program in the Cervus3.0 software to carry out data analysis. The LOD value in the table is the logarithmic value of the paternity index (paternity index). If the LOD value is greater than 0, it means that compared with any individual, the candidate parent (Candidate Parent) is most likely to be the real parent; LOD value less than 0 means that compared with any individual, the candidate parent cannot be the rea...
Embodiment 3
[0072] Example 3 Application of 8 microsatellite loci in identification of genetic relationship in dairy cows
[0073] In this example, a pair of parent-offspring with a clear parent-child relationship and an unrelated parent were selected from the dairy cow population for verification, simulating parentage identification under normal production conditions.
[0074] sample
[0075] (1) 13325 blood samples of offspring cows, No. 1;
[0076] (2) Frozen semen of candidate male bull 37310001, No. 2;
[0077] (3) Frozen semen of irrelevant male bull 37310026, number 3.
[0078] Identification requirements: DNA test, paternity test
[0079] Test and Results
[0080] Extract the DNA of the above-mentioned sample to be tested, and perform PCR amplification on 8 microsatellite loci;
[0081] Taking the 15 μl amplification system as an example, the reaction system for PCR amplification is shown in Table 2.
[0082] Table 2 PCR amplification system
[0083]
[0084] The PCR rea...
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